Project Summary/Abstract There is a paradigm shift in testing and modeling of chemical toxicity. Current toxicology calls for human-cell- based (vs. animal-based), evidence-based, and Omics-based assays. Mesenchymal stem cells (MSC) play important roles in the maintenance and repair of various tissues, including bone, cartilage, and muscle. MSCs are also found in circulating blood and participate in the wound-healing process. Therefore, MSC is arguably one of the most physiologically relevant adult stem cells for chemical toxicology. In response to RFA-ES-13- 003, Newomics Inc. proposes to develop high-throughput Omics-based assays to evaluate the effects of chemical toxicants on proliferation and differentiation of human MSC. The core technology platform will be based on Newomics' breakthrough silicon-microfluidic-chip, the multinozzle emitter array chip (MEA chip), which enables nanoflow liquid chromatography-electrospray ionization mass spectrometry (nanoLC-ESI/MS)- based, high-sensitivity, high-throughput, and multiplex measurements of multiclass analytes (peptides, proteins, and metabolites) at the Omics level, from small volumes of samples. The project will also take advantage of our extensive experience in the characterization of human MSC and mechanistic studies of their responses to genotoxic stresses such as ionizing radiation and chemical toxicants such as hydrogen peroxide. In this Phase I project, we will demonstrate a robust and universal two-stage workflow for Omics-based and high-content screening of chemical toxicity using human MSC. In Aim 1, we will develop phenotypic assays to prioritize the conditions for the focused follow-up Omics-based assays. In Aim 2, we will perform high- throughput proteomic and metabolomic analyses of MSC before and after chemical treatments, and the associated medium supernatants from each well on the 96-well or 384-well plates used for the quantitative high-throughput screening (qHTS). We will perform bioinformatics analysis and delineate the pathways and networks responsible for the chemical toxicity to MSC. If successful, Newomics' MEA chip may become a universal platform for Omics-based chemical toxicology using stem cells.